Small conductance Ca-activated K channels (SK channels) affect excitability and contribute to shaping excitatory postsynaptic potentials (EPSPs). SK2 channels are expressed throughout the dendritic arbors of hippocampal CA1 neurons and contribute to synaptic plasticity and learning and memory. We have recently found that the SK2 gene encodes two isoforms of the SK2 protein. SK2-L contains an additional 209 N- terminal amino acids compared to SK2-S that is completely contained within the SK2-L protein. In mice that selectively lack the SK2-L isoform (SK2-Sonly) dendritic and spine expression is drastically reduced and the SK2 channel contribution to synaptically evoked EPSPs is abolished. These results form the basis for the overriding hypothesis of this application: The unique SK2-L N-terminal domain confers dendritic and spine localization and function to SK2 channels that is accomplished by selective association with targeting proteins. We will use an integrated repertoire of electrophysiology, immunoEM, molecular biology, and biochemistry to test the following specific hypotheses. 1. Hypothesis: The loss of either isoform alters CA1 cellular physiology and hippocampal-dependent learning. Use SK2-Sonly and SK2-Lonly mice to determine: A) the subcellular distributions of SK2-L and SK2-S in CA1 neurons; B) the effects of SK2-S and SK2-L on synaptic plasticity and EPSPs, and on SK2 internalization following the induction of LTP; C) memory encoding in hippocampal-dependent spatial and non-spatial learning tasks. WT, SK2-Sonly/SK2-Lonly, and SK2-/- mice will serve as controls. 2. Hypothesis: The SK2-L N-terminus contains determinants that are necessary and sufficient for dendritic targeting. A) Determine the subcellular distributions of SK2-S, or SK2-L without or with SK2-S expressed in hippocampal neurons from SK2-/- mice that lack both SK2 isoforms. B) i. Identify the domains responsible for dendritic localization; ii. Determine whether these motifs are autonomous. 3. Hypothesis: Proteomics using SK2-Lonly or SK2-Sonly mice will identify proteins that specifically co-assemble with SK2-L subunits and mediate dendritic targeting. Protein complexes associated with SK2 channels will be immunopurified from SK2-Lonly or SK2-Sonly mouse brain using an antibody specific for SK2-L, or a pan-SK2 antibody, respectively. Parallel experiments using SK2-/- mice will serve as a negative control. Proteins that specifically co-assemble with SK2-L will be identified by mass spectrometry and investigated for their functional significance. PUBLIC HEALTH RELEVANCE: SK2 channels, one type of Ca2+-activated K+ channel, influence learning and memory. Blocking SK2 channels facilitates learning in animal models of brain damage, and reverses the navigation failure in an animal model associated with the development of cognitive disorders in Alzheimer's disease as well as during normal brain aging. These studies will reveal novel mechanisms for how SK2 channels influence information processing and storage in the brain, and will suggest novel interventional strategies that target different isoforms of SK2 channels to treat learning deficits associated with trauma, pathology and normal aging.